Our research project will focus on the exploration of unresolved problems concerning the regulation of renal potassium (K+) channels (ROMK). The kidney plays a major role in potassium homeostasis by a balance between potassium secretion in principal tubule cells and potassium reabsorption in intercalated cells of the distal nephron. Following active uptake of potassium across the basolateral membrane by Na-K+ ATPase, diffusion across the apical membrane of principal cells occurs via low-conductance potassium channels. These channels have been cloned and we propose to explore the mechanism of their regulation. Two major goals will be pursued. First, we propose to explore the regulation of nucleotidedependent channel gating by kinases, pH and phospolipids, and the stoichiometry of channel subunits. Second, because recent studies have identified ROMK deletions in Bartter's syndrome, a condition of renal sodium (Na) and K+ wasting, we plan to study the effects of loss of apical potassium channels in a ROMK knock-but model. We will also examine alternative transport mechanisms of K+ secretion. Patch-clamp and whole-cell current measurements will be carried out in native tubules and in an oocyte expression system, and tubule perfusion studies in ROMK knock-out mice.